title: EGFR mutation detection by microfluidic technology: a validation study.
abstract: Advanced non-small cell lung cancer samples are tested for epidermal growth factor receptor (EGFR) gene mutations. Their detection by direct sequencing is time-consuming. Conversely, the length analysis of fluorescently labelled PCR products is easier. To avoid labelled primers and the automated capillary electrophoresis apparatus, we validated a fast and sensitive chip-based microfluidic technology. The limit of detection of fragment length assay on microfluidic device was 5%, more sensitive than direct sequencing (12.5%). The novel methodology showed high accuracy in the analysis of samples whose mutational status was known. The accuracy in quantifying mutated alleles (mA) was evaluated by PCR products subcloning; the mA% provided by direct sequencing of subcloned PCR products showed a close correlation with the mA% provided by the microfluidic technology for both exon 19 (R(2)=0.9) and 21 (R(2)=0.9). Microfluidic-based on-chip electrophoresis makes EGFR testing more rapid, sensitive and cost-effective.
authors.affiliation: Dipartimento di Sanità Pubblica, Università di Napoli Federico II, , Napoli, Italia.
authors.forename: Umberto
authors.initials: U
authors.lastname: Malapelle
authors.forename: Stefania
authors.initials: S
authors.lastname: Russo
authors.forename: Francesco
authors.initials: F
authors.lastname: Pepe
authors.forename: Roberta
authors.initials: R
authors.lastname: Sgariglia
authors.forename: Caterina
authors.initials: C
authors.lastname: De Luca
authors.forename: Claudio
authors.initials: C
authors.lastname: Bellevicine
authors.forename: Pierlorenzo
authors.initials: P
authors.lastname: Pallante
authors.forename: Giancarlo
authors.initials: G
authors.lastname: Troncone
mesh_terms: D000230:Adenocarcinoma / Q000235:genetics*; D000483:Alleles; D002289:Carcinoma, Non-Small-Cell Lung / Q000235:genetics*; D002460:Cell Line; D003362:Cost-Benefit Analysis; D004252:DNA Mutational Analysis / Q000379:methods; D004273:DNA, Neoplasm / Q000737:chemistry / Q000235:genetics; D019075:Electrophoresis, Capillary / Q000379:methods; D046209:Electrophoresis, Microchip / Q000379:methods; D066246:ErbB Receptors / Q000235:genetics*; D005091:Exons / Q000235:genetics; D006801:Humans; D057230:Limit of Detection; D008175:Lung Neoplasms / Q000235:genetics*; D046210:Microfluidic Analytical Techniques / Q000379:methods*; D009154:Mutation; D012150:Polymorphism, Restriction Fragment Length; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA; D013997:Time Factors
chemical_list: D004273:DNA, Neoplasm; C512478:EGFR protein, human; D066246:ErbB Receptors
keywords: EGFR; LUNG; MOLECULAR PATHOLOGY